Pituitary tumors are the most common brain tumor, found at autopsy in up to 20% of our population and clinically in 1/10,000 persons. With the current number of US veterans, this translates into over 225,000 veterans with pituitary tumors. Gonadotrope (or null cell) tumors occur more commonly in men who present clinically with erectile dysfunction, hormone deficiencies, headaches and visual disturbances progressing to blindness. Many of these tumors are invasive and recur. There are no available medical therapies for these large pituitary tumors and surgery remains the treatment of choice. The genetic and molecular mechanisms underlying pituitary tumorigenesis are poorly understood. We have amassed a unique bank of >300 human pituitary tumor and 100 normal pituitary samples tied to a clinical database. We performed a combined genetic screen to identify copy number alternations together with genomic transcriptome profiling using microarrays. Amplification of the MST4 gene, a Sterile 20-like germinal center kinase (GCK) III subfamily member was detected in an aggressive gonadotrope pituitary tumor together with increased MST4 transcript and protein levels in all pituitary tumors. Extensive new preliminary data demonstrate the role of MST4 driving proliferation and survival in response to oxidative stress and hypoxia to promote tumorigenesis. In this revised Merit Review we propose: Aim 1: To determine the frequency of altered copy number, translocation or mutation in the MST4 kinase as a mechanism contributing to human pituitary tumor pathogenesis. These experiments will define the mechanism of MST4 dysregulation in human pituitary tumors. Aim 2: To examine the importance of the kinase domain of MST4 to mediate its effects on proliferation and survival and confirm or refute the role of MST4 to mediate tumor growth in a preclinical mouse model. Studies using gonadotrope and growth hormone pituitary cells based systems will dissect the critical regions of MST4 to mediate tumorigenesis using read outs of proliferation, survival in response to oxidative stress or hypoxia and clonagenic assays. Results will be confirmed in a preclinical ex vivo nude mouse model to confirm the ability to MST4 to promote tumorigenesis in vivo. Aim 3: To identify small molecules with the capacity to inhibit MST4 activity and establish the structural characteristics necessary for selective MST4 inhibition. Library screening of known compounds together with in silico computer screening will identify known or design new candidate MST4 kinase inhibitors that will be tested in the cell and animal models as proof of concept for our ultimate goal of targeting MST4 in human pituitary tumors. Together these studies will use cutting edge genetic and genomic approaches together with mechanistic studies to dissect the role of dysregulation of MST4 in human pituitary tumors. These studies will shed light into the pathobiology of pituitary tumors. Targeting of the MST4 kinase may provide novel medical treatment strategies for our patients with pituitary tumors and other malignancies where MST4 is dysregulated.